This research project includes an investigation of the physiological mechanisms regulating the activation and control of human complement. We will investigate the C3b- and C4b- mediated inhibition of immune complex-inducer C1 activation. The effects of factors H, I and C4b-binding protein on this feedback inhibition will be assessed. We will determine if the covalent binding of nascent C3b and C4b to antibody, antigen or other complement proteins is crucial. The effect of antibody subclass on the strength of feedback inhibition will be studied. Are nonimmune activators of C1 also inhibited by nascent C3b and C4b? We will also determine if feedback inhibition correlates with antibody-antigen dissociation. The inhibiting properties of the C4A isotype of C4 will be compared with the C4B isotype in order to gain insight as to the chemical group to which the inhibiting molecules bind. (C4A prefers binding to amino groups, while C4B prefers hydroxyl groups). If, as preliminary data suggest, immunoglobulin is the receptor molecule for the inhibiting complement protein(s), we will then selectively modify different amino acids (or carbohydrate) of the immunoglobulin. The goal is to block the C3b and/or C4b binding site(s) without affecting the complementing activating or antigen binding activities. (If C1q turns out to be the receptor molecule, it will be similarly treated.) An antibody that is successfully modified (i.e., so that it is not subject to feedback inhibition by C3b and C4b) would be a potent complement activator, able to generate high levels of biologically active mediators upon binding to antigen. This modified antibody might be of great practical use for monoclonal antibody therapy. We will also study the effects of cellular-derived protease inhibitors (termed protease nexin or PN) on complement activation. PN-I will be isolated from cultured fibroblasts and characterized as to its ability to inhibit activated C1, the fluid phase activation of C1 and immune complex-induced activation of C1. Comparisons will be made with the plasma protein C1- inhibitor. Furthermore, quantitative and qualitative comparisons will be made between fibroblasts derived from normal individuals vs. those with hereditary angioedema (HAE), a disease characterized by a genetic deficiency in C1-inhibitor function. Since the pathological processes leading to edema may be also occurring outside the blood vessel, these studies are relevant for a better understanding of the disease.